All jawed vertebrates so far examined carry both alpha/beta and gamma/delta TCR genes, suggesting that both T cells types are present and important to survival. Although many animals have approximately equal numbers of gamma/delta and alpha/beta T cells, in the lymphoid organs of both mice and humans, the gamma/delta T cells are much less numerous than are alpha/beta T cells. There are a host of additional similarities between the mouse and human immune systems, so perhaps in both, gamma/delta T cells have been reduced to their most essential functions. Gamma/delta T cells have been shown to be important in clearing infectious agents, in the destruction of tumor cells, and in controlling immune and inflammatory responses. In this study, we will examine what causes a high frequency of female mice of the C57Bl/10 strain lacking gamma/delta T cells (B10.TCRdelta-/-) to develop inflammation of the cornea. The fact that keratitis arises spontaneously in these mice suggests that gamma/delta T cells are normally needed to prevent an immune attack on the eye. Our preliminary data indicate that keratitis in these mice is not infectious, but is instead autoimmune, and is mediated by auto aggressive ab T cells. Also, we showed that adoptively transferring a particular gamma/delta T cell subset reduces the incidence of keratitis. In this proposal, we will test the hypothesis that immunoregulatory gamma/delta T cells normally prevent cornea-specific autoreactive alpha/beta T cells from attacking the cornea. Thus, B10.TCRdelta-/- mice would be susceptible to keratitis because these regulatory gamma/delta T cells are missing. The specific aims of this project are: Specific Aim 1 - to test the hypothesis that keratitis-inducing alpha/beta T cells in B10.TCRdelta-/-mice are cornea-specific auto aggressive cells. We will investigate the characteristics of alpha/beta T cells that infiltrate the corneas of keratitic B10.TCRdelta-/- mice, examine the cytokines they produce when activated, and test whether they respond in vitro to corneal antigen. We will also determine which alpha/beta T cells are necessary to transfer the disease, and by intravital imaging whether they directly attack cells in the cornea. Specific Aim 2- to test the hypothesis that gamma/delta T cells which protect against keratitis tolerize disease-inducing alpha/beta cells. We will characterize a population of gamma/delta T cells that normally resides in the cornea, and examine whether splenic gamma/delta T cells that reduce the incidence of keratitis contain similar cells. We will also test whether gamma/delta T cells carry out their protective rol while in the cornea, determine which of the cytokines they produce are critical for their effect. Whether they act directly on alpha/beta T cells will be examined by intravital imaging of the cornea. We will also investigate their ability to induce regulatory alpha/beta T cells, and as well as to promote healing of the cornea during keratitis. This project is both innovative and relevant because it focuses on a new small animal model for spontaneous autoimmune keratitis which lacks the drawbacks and limitations of existing small animal models for this disease. Gamma/delta T cells have already been implicated by others in human eye diseases. This project could therefore lead to improvements or refinements in the treatment of corneal disease in humans, and will contribute towards our understanding of immune privilege in the eye. In addition, its findings are likely to reveal unique attributes of gamma/delta T cells that may help o explain why they have been evolutionarily conserved. PUBLIC HEALTH RELEVANCE: We recently discovered that female mice of a certain background, when unable to produce a rare type of T cell known as gamma/delta T lymphocytes, spontaneously develop an inflammation of the cornea of the eye, or keratitis, at a very high rate. This disease, which can become very severe, appears to stem from an autoimmune attack by conventional T cells, or alpha/beta T cells, on the cornea. We hypothesize that gamma/delta T cells normally suppress these autoimmune alpha beta T cells and prevent keratitis. We believe these mice represent a useful disease model for autoimmune keratitis, and propose to further investigate the cells that are responsible for and those that prevent this disease.